Normal growth control is dependent upon the coordinate expression of diverse genes that act to constrain proliferation within appropriate limits. Regulation of these genes is mediated by multimeric factors that either promote or repress transcription. However, the ability of transcription factors (TFs) to form the active unit is contingent upon strict conformational requirements, among which is the establishment of appropriate sulfhydryl interactions. Consistent with this paradigm is the finding by the investigators laboratory and others that the binding of some TFs to their cognate DNA recognition sites is highly sensitive to the redox environment. The investigator and colleagues have also observed that the expression of the gene for the sulfhydryl- oxidoreductase protein disulfide isomerase (PDI) and its associated chaperone-like enzymatic activity are modulated with the cell cycle. Moreover, PDI is capable of influencing the in vitro responsiveness of NF-kB and fos-jun activity to the redox state. Collectively, these results suggest that a coupled redox-chaperone signalling pathway may exist whereby localized changes in redox potential and/or thio-modifying chaperone activity can be functionally relevant in regulating genes critical to the control of cellular proliferation. The research described in this proposal is directed towards assessing this hypothesis by (1) determining whether changes in cellular redox state are associated with changes in the proliferative state of the cell; (2) determining the causal relationship, if any, between the redox environment, PDI and other thio-modifying chaperones, and the proliferative capacity of the cell; (3) determining whether any induced changes in redox environment and PDI/chaperone activity alters the in vivo expression of genes regulated by NF- kB and fos-jun that have been implicated as being involved in growth control. Changes in redox states will be measured by determining the intracellular concentrations of molecules that are the principal determinants of the redox potential. The causal relationship will be studied by characterizing the effect on cellular proliferation of chemicals that disrupt normal glutathione metabolism and thereby modulate the redox potential. This would be done in conjunction with the use of specific inhibitors that disrupt the action of thio-modifying enzymes. The effect of redox and chaperone activity on expression of endogenous and recombinant reporter genes will be studied by northern analysis and RT-PCR. By using both normal and malignant fibroblast cells which differ in the stringency of their growth control, special emphasis will be placed on extrapolating the results of this study to the possible role of a redox/chaperone couple defect in the oncogenic state. Central to the proposed research is the involvement of both undergraduate and graduate students. This investigation provides an opportunity for integrating undergraduate students into the research effort by interfacing with the special Freshman Scholars Research Program and other undergraduate research options afforded students at the Catholic University.